Performance evaluation of two antigen-extracted xenogeneic ostein and experimental study on repairing skull defects in rats

Abstract

To evaluate the physical and chemical properties, immunogenicity, and osteogenesis of two antigen-extracted xenogeneic bone scaffolds-decalcified bone matrix (DBM) and calcined bone. By removing the inorganic and organic components of adult pig femus, xenogeneic DBM and calcined bone were prepared respectively. The density and pH value of the two materials were measured and calculated, the material morphology and pore diameter were observed by scanning electron microscope, and the surface contact angle was measured by automatic contact angle measuring instrument. The safety, osteogenic activity, and immunogenicity of the two materials were evaluated by cytotoxicity test, osteoblast proliferation test, DNA residue test, and human peripheral blood lymphocyte proliferation test. The two materials were implanted into the 5 mm full-thickness skull defect of 6-week-old male Sprague Dawley rats (the blank control group was not implanted with materials). The materials were taken at 4 and 8 weeks after operation, the repair effect of the materials on the rat skull was observed and evaluated by gross observation, Micro-CT scanning, and HE staining observation. Compared with calcined bone, DBM has lower density and poor hydrophilicity; the pH value of the two materials was 5.5-6.1, and the pore diameter was 160-800 μm. The two materials were non-cytotoxic and could promote the proliferation of osteoblasts. The absorbance ( A) values of osteoblast proliferation at 1, 4, and 7 days in the DBM group were significantly higher than those in the calcined bone group ( P<0.05). The DNA residues of the two materials were much lower than 50 ng/mg dry weight, and neither of them could stimulate the proliferation and differentiation of human peripheral blood lymphocytes. The results of animal experiments in vivo showed that the bone volume/total volume (BV/TV) in DBM group and calcined bone group were significantly higher than that in blank control group at 4 weeks after operation ( P<0.05), and that in calcined bone group was significantly higher than that in DBM group ( P<0.05); at 8 weeks after operation, there was no significant difference in BV/TV between groups ( P>0.05). HE staining showed that at 4 and 8 weeks after operation, the defect in the blank control group was filled with fibrous connective tissue, the defect was obvious, and no bone growth was found; the defect in DBM group and calcined bone group had been repaired to varying degrees, and a large number of new bone formation could be seen. The material degradability of DBM group was better than that of calcined bone group. The physical and chemical properties and degradability of the two kinds of xenogeneic bone scaffolds were slightly different, both of them have no immunogenicity and can promote the repair and reconstruction of skull defects in rats.